Journal: Oncotarget

Article Title: A novel microfluidic model can mimic organ-specific metastasis of circulating tumor cells

doi: 10.18632/oncotarget.9382

Figure Lengend Snippet: ( A – C ) Typical metastatic colonies of MCF-7, MDA-MB-231, and ACC-M cells after HE staining and anti-human Pan CK immunostaining in the lung of nude mice. The metastatic colonies are circumscribed by black dotted lines and indicated by asterisk. Scale bar = 50 μm. ( D ) The metastasis rates and the colony numbers of MCF7, MDA-MB-231, and ACC-M, respectively. ( E ) The metastasic colony areas of MCF-7, MDA-MB-231, and ACC-M cells. The colony areas of MDA-MB-231 cells were significantly higher than those of MCF-7 cells (** p < 0.01, n = 5 per group).

Article Snippet: Immunohistochamical staining with anti-humna Pan CK (Merckmillipore, USA) was performed to detect the metastatic colonies.

Techniques: Staining, Immunostaining

Journal: International Journal of Molecular Sciences

Article Title: Role of HMGB1 in an Animal Model of Vascular Cognitive Impairment Induced by Chronic Cerebral Hypoperfusion

doi: 10.3390/ijms21062176

Figure Lengend Snippet: Increased high-mobility group box protein 1 (HMGB1) and its proinflammatory cytokines (tumor necrosis factor-alpha [TNF-α] and interleukin [IL]-1β) in the cortex and hippocampus at 3 months after chronic cerebral hypoperfusion (CCH). ( A ) Western blot results of HMGB1 in the cortex and hippocampus. ( B ) Quantification results of HMGB1 relative protein in the cortex and hippocampus. ( C ) Immunostaining of HMGB1, TNF-α, and IL-1β in the cortex. ( D ) Quantification data of immunostaining image. n = 6 for each group. Data are presented as the mean + standard deviation; * p < 0.05; ** p < 0.01 with unpaired t-test; scale bar = 50 μm.

Article Snippet: Equal amounts of protein (20 μg) were then separated through sodium dodecyl sulfate–polyacrylamide gel electrophoresis and transferred to a polyvinylidene difluoride membrane, which was subsequently incubated in a primary Ab against HMGB1 (mouse monoclonal Ab, 1:1000, GeneTex, Irvine, CA, USA), interleukin (IL)-1α (rabbit polyclonal Ab, 1:1000, GeneTex, Irvine, CA, USA), tumor necrosis factor-alpha (TNF-α; rabbit polyclonal Ab, 1:1000, GeneTex, USA), and IL-6 (rabbit polyclonal Ab, 1:1000, GeneTex, Irvine, CA, USA).

Techniques: Western Blot, Immunostaining, Standard Deviation

Journal: International Journal of Molecular Sciences

Article Title: Role of HMGB1 in an Animal Model of Vascular Cognitive Impairment Induced by Chronic Cerebral Hypoperfusion

doi: 10.3390/ijms21062176

Figure Lengend Snippet: The effect of high-mobility group box protein 1 (HMGB1) suppression with CRISPR/Cas9 knockout (KO) plasmid on proinflammatory cytokines, hippocampus volume, and memory function. ( A ) Decreased level of tumor necrosis factor-alpha, interleukin (IL)-1β, and IL-6 of bilateral common carotid artery occlusion (BCCAO) HMGB1-KO mice showed with immunostaining. ( B ) Attenuation of hippocampal atrophy in BCCAO HMGB1-KO mice but not in BCCAO controls (the area of the hippocampus is indicated with a red circle). ( C ) Improvement of memory decline in BCCAO HMGB1-KO mice but not in BCCAO controls. n = 5 for each group. Data are presented as the mean + standard deviation; * p < 0.05; ** p < 0.01; *** p < 0.001 with one-way analysis of variance (ANOVA) followed by Sidak’s multiple comparison test; novel object recognition test before and after CRISPR/Cas9 injection with two-way ANOVA (F(3,24) = 7.598, p < 0.01); scale bar = 100 μm.

Article Snippet: Equal amounts of protein (20 μg) were then separated through sodium dodecyl sulfate–polyacrylamide gel electrophoresis and transferred to a polyvinylidene difluoride membrane, which was subsequently incubated in a primary Ab against HMGB1 (mouse monoclonal Ab, 1:1000, GeneTex, Irvine, CA, USA), interleukin (IL)-1α (rabbit polyclonal Ab, 1:1000, GeneTex, Irvine, CA, USA), tumor necrosis factor-alpha (TNF-α; rabbit polyclonal Ab, 1:1000, GeneTex, USA), and IL-6 (rabbit polyclonal Ab, 1:1000, GeneTex, Irvine, CA, USA).

Techniques: CRISPR, Knock-Out, Plasmid Preparation, Immunostaining, Standard Deviation, Comparison, Injection

Journal: PLoS ONE

Article Title: Glycoprotein Nonmetastatic Melanoma B (Gpnmb)-Positive Macrophages Contribute to the Balance between Fibrosis and Fibrolysis during the Repair of Acute Liver Injury in Mice

doi: 10.1371/journal.pone.0143413

Figure Lengend Snippet: (A) Hepatic expression of IL-1β, IL-10, and CCL2 are significantly decreased (clodronate liposome injection group [ n = 3, only survival] vs. PBS injection group [ n = 4)]). Values are mean ± SEM. * P < 0.05 (Mann-Whitney U test)). (B) Collagen deposition and α-SMA expression are decreased (original magnification, x400). (C) Hepatic expression of TGF-β, Col1α1, and MMP-13 are significantly decreased (clodronate liposome injection group [ n = 3, only survival] vs. PBS injection group [ n = 4]). Values are mean ± SEM. * P < 0.05 (Mann-Whitney U test)).

Article Snippet: For immunohistochemical analysis, liver specimens were fixed in 10% buffered formalin or 4% paraformaldehyde and incubated with rat anti-mouse F4/80 (Clone: A3-1, AbD Serotec, Raleigh, NC, USA), rat anti-mouse CD68 (Clone: FA-11, AbD Serotec), goat anti-mouse Gpnmb (Clone: #297310, R&D Systems, Minneapolis, MN, USA), and monoclonal antibody against α-SMA (Clone: E184, Merck Millipore, Billerica, MA, USA).

Techniques: Expressing, Injection, MANN-WHITNEY

Journal: PLoS ONE

Article Title: Glycoprotein Nonmetastatic Melanoma B (Gpnmb)-Positive Macrophages Contribute to the Balance between Fibrosis and Fibrolysis during the Repair of Acute Liver Injury in Mice

doi: 10.1371/journal.pone.0143413

Figure Lengend Snippet: (A) Expression of Gpnmb is observed in DBA-g+ but not DBA mice following a single injection of CCl 4 . However, (B) sequential changes in serum ALT levels and the degree of liver injury are not affected by the lack of Gpnmb-positive macrophages (original magnification, x100). Conversely, (C) the areas of fibrosis and (D) the number of α-SMA-positive cells are significantly decreased in the liver tissues of DBA mice compared to DBA-g+ mice (original magnification, x200). Additionally, (E) although lack of Gpnmb-positive macrophages does not affect expression of TGF-β or Col1α1, expression of MMP-9, MMP-13, and TIMP-1 are significantly decreased in mice lacking Gpnmb expression at six or eight days after single injection of CCl 4 . Values are mean ± SEM ( n = 4). * P < 0.05 (Mann-Whitney U test).

Article Snippet: For immunohistochemical analysis, liver specimens were fixed in 10% buffered formalin or 4% paraformaldehyde and incubated with rat anti-mouse F4/80 (Clone: A3-1, AbD Serotec, Raleigh, NC, USA), rat anti-mouse CD68 (Clone: FA-11, AbD Serotec), goat anti-mouse Gpnmb (Clone: #297310, R&D Systems, Minneapolis, MN, USA), and monoclonal antibody against α-SMA (Clone: E184, Merck Millipore, Billerica, MA, USA).

Techniques: Expressing, Injection, MANN-WHITNEY

Journal: Open Biology

Article Title: Neuroanatomical characterization of the Nmu-Cre knock-in mice reveals an interconnected network of unique neuropeptidergic cells

doi: 10.1098/rsob.220353

Figure Lengend Snippet: Primary antibodies.

Article Snippet: anti-estrogen receptor alpha (ER α ) , polyclonal , rabbit , Merck Millipore, USA (06-935) , 1 : 1000 , AB_310305.

Techniques:

Journal: Scientific Reports

Article Title: Viral-like TLR3 induction of cytokine networks and α-synuclein are reduced by complement C3 blockade in mouse brain

doi: 10.1038/s41598-023-41240-z

Figure Lengend Snippet: Intra-striatal TLR3 activation in mouse brain increased complement C3 and α-synuclein protein levels. WT mice received a single unilateral deposit of 50 μg Poly(I:C) (n = 10) or vehicle (n = 9) in the striatum. Two days later, injected striata were isolated and homogenised in RIPA buffer for protein detection by Western blot ( A, E ). ( B – D, F – I ) Histogram bars represent the mean levels of target protein normalized to GAPDH or total protein visualized by stain-free UV-fluorescence. Uncropped blots can be found in Figs. – . Error bars represent SEM. A.U., arbitrary units. Statistical analysis was performed using unpaired t-tests. ns, p > 0.05; *, p < 0.05; ***, p < 0.001.

Article Snippet: Membranes were blocked for 30 min with 5% milk in Tris-buffered saline containing 0.1% Tween 20 and were probed overnight at 4 °C with primary antibodies against the following target proteins: α-synuclein (BD biosciences, BD610787 1:2000), C3 (Abcam, ab200999, 1:1000), C1q (Abcam, ab182451, 1:500), Iba1 (Abcam, ab178846, 1:1000), GFAP (EMD Millipore, MAB360, 1:2000), GAPDH (Cell Signalling Technology, #2118, 1:2000).

Techniques: Activation Assay, Injection, Isolation, Western Blot, Staining, Fluorescence

Journal: Scientific Reports

Article Title: Viral-like TLR3 induction of cytokine networks and α-synuclein are reduced by complement C3 blockade in mouse brain

doi: 10.1038/s41598-023-41240-z

Figure Lengend Snippet: Knockdown of C3 prior to TLR3 activation reduced α-synuclein protein levels in the striatum of mouse brain. Alpha-synuclein levels in the injected striatum of mice from NT ASO/Poly(I:C) (n = 10) and C3 ASO/Poly(I:C) (n = 10) groups were measured by Western blot ( A ). The uncropped blot can be found in Fig. . ( B ) Histogram bars represent the mean levels of α-synuclein normalized to total protein loading. Error bars represent SEM. A.U., arbitrary units. Statistical analysis was performed using unpaired t-tests. ***, p < 0.001.

Article Snippet: Membranes were blocked for 30 min with 5% milk in Tris-buffered saline containing 0.1% Tween 20 and were probed overnight at 4 °C with primary antibodies against the following target proteins: α-synuclein (BD biosciences, BD610787 1:2000), C3 (Abcam, ab200999, 1:1000), C1q (Abcam, ab182451, 1:500), Iba1 (Abcam, ab178846, 1:1000), GFAP (EMD Millipore, MAB360, 1:2000), GAPDH (Cell Signalling Technology, #2118, 1:2000).

Techniques: Activation Assay, Injection, Western Blot